The Section of Transgenesis (TS) in Laboratory of Neurogenetics (LNG) was devoted to study the pathogenic mechanisms and experimental therapeutics of neurodegenerative diseases by a combination of in vivo mouse modeling and in vitro neurobiology approaches. Our research covers three major neurological diseases: Alzheimers disease (AD), Parkinsons disease (PD), and Amyotrophic lateral sclerosis (ALS). [unreadable] [unreadable] Project 1. BACE1 is Critical for Cognition [unreadable] A transmembrane aspartyl protease termed beta-site APP cleavage enzyme1 (BACE1) that cleaves the amyloid precursor protein (APP) is required for the generation of amyloid-beta (A-beta) peptides implicated in the pathogenesis of AD. Significantly, deletion of BACE1 in a mouse model of AD, the APPswe; PS1delE9 double transgenic mice prevents both A-beta deposition and age-associated cognitive abnormalities that occur in this model of A-beta amyloidosis. Moreover, the A-beta burden is sensitive to BACE1 dosage in young but not in aged APPswe; PS1delE9 mice, suggesting that A-beta clearance mechanisms in aged animals may be compromised. Although BACE1 null mice do not exhibit overt developmental abnormalities or adult-onset neuropathology, however, these animals do manifest alterations in performance on tests of cognition and emotion. Importantly, the memory deficits occurring in BACE1 knockout mice are prevented in APPswe; PS1delE9; BACE1knockout mice. Our results establish that BACE1-dependent APP processing is critical for cognitive and emotional behaviors, suggesting that future studies should be alert to potential mechanism-based toxicities associated with BACE1 inhibitors designed to ameliorate A-beta amyloidosis in AD. These findings were published in the Journal of Neuroscience last year.[unreadable] [unreadable] Project 2. Activation of protein kinase C modulates BACE1-mediated b-secretase activity[unreadable] Previous studies suggest that activation of protein kinase C (PKC) modulates the b-secretase-mediated cleavage of APP and reduces the production of Ab. The mechanism of PKC-mediated modulation of b-secretase activity, however, remains elusive. We report here that activation of PKC modulated b-secretase activity through either suppressing the accumulation or promoting the translocation of BACE1 protein in a cell type-dependent manner. We found that activation of PKC suppressed the accumulation of BACE1 protein in fibroblasts through an enhancement of intracellular protease activities. In neurons, activation of PKC did not alter the expression level of BACE1, but led to more BACE1 translocated to the cell surface, resulting in a decreased cleavage of APP at the b1 site. Together, Our findings provide novel mechanisms of PKC-mediated modulation of b-secretase activity, suggesting that alteration of the intracellular trafficking of BACE1 may serve as a useful therapeutic strategy to lower the production of Ab in AD. This work is under revision from Neurobiology of Aging. [unreadable] [unreadable] Project 3. Loss Function of Alsin is Not Sufficient to Trigger Major Motor Neuron Degeneration but Predispose Neurons to Oxidative Stress[unreadable] ALS, the most common motor neuron disease, is caused by a selective loss of motor neurons in the central nervous system. Mutations in the ALS2 gene have been linked to one form of autosomal recessive juvenile onset ALS (ALS2). To investigate the pathogenic mechanisms of ALS2, we generated ALS2 knockout (ALS2-/-) mice. While ALS2-/- mice lacked obvious developmental abnormalities, they exhibited age-dependent deficits in motor coordination and motor learning. Moreover, ALS2-/- mice showed a higher anxiety response in the open field and elevated plus maze tasks. Although they failed to recapitulate clinical or neuropathological phenotypesconsistent with motor neuron disease by 20 months of age, ALS2-/- mice or primary cultured neurons derived from these mice were more susceptible to oxidative stress compared to wild type controls. These observations suggest that loss of ALS2 function is insufficient to cause major motor deficits or motor neuron degeneration in a mouse model, but predisposes neurons to oxidative stress. These data were published in Journal of Neuroscience this August.[unreadable] [unreadable] Project 4. ALS2-Deficiency Leads to Neuronal Degeneration in Amyotrophic Lateral Sclerosis through Altered AMPA Receptor Trafficking[unreadable] To further understand the function of alsin that is encoded by the full-length ALS2 gene, we screened proteins interacting with alsin. Here we report that alsin interacted with glutamate receptor interacting protein 1 (GRIP1) both in vitro and in vivo, and co-localized with GRIP1 in neurons. In support of the physiological interaction between alsin and GRIP1, the subcellular distribution of GRIP1 was altered in ALS2-/- spinal motor neurons, which correlates with a significant reduction of a-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) type glutamate receptor subunit 2 (GluR2) at the synaptic/cell surface of ALS2-/- neurons. The decrease of calcium-impermeable GluR2-containing AMPA receptors at the cell/synaptic surface rendered ALS2-/- neurons more susceptible to glutamate receptor-mediated neurotoxicity. Our findings reveal a novel function of alsin in AMPA receptor trafficking and provide a novel pathogenic link between ALS2-deficiency and motor neuron degeneration, suggesting a protective role of alsin in maintaining the survival of motor neurons. This work is under review from Journal of Neuroscience.[unreadable] Project 5. Deficiency in ALS2 gene does not affect the motor neuron degeneration in SOD1G93A transgenic mice [unreadable] Previous in vitro studies suggest that over-expression of ALS2 protects cells from mutant Cu/Zn superoxide dismutase (SOD1)-induced cytotoxicity. To test whether ALS2 plays a protective role against mutant SOD1-mediated motor neuron degeneration in vivo, we examined the progression of motor neuron disease in SOD1G93A mice on an ALS2 null background. Our data suggest that deficiency in the ALS2 gene does not affect the pathogenesis of SOD1G93A mice. These data were published in Neurobiology of Aging this September.[unreadable] [unreadable] Project 6. A Novel Function of Alsin in Regulating Endosome Degradation that is up-regulated in a Mouse Model of Amyotrophic Lateral Sclerosis [unreadable] Alsin is a newly defined guanine-nucleotide-exchange factor of Rab5 family small GTPase. Dysfunction of alsin has been linked to one form of juvenile onset recessive familial ALS (ALS2). However, how loss of alsin affects Rab5-mediated endocytosis is unclear. Here we report that the Rab5-mediated endosome trafficking is severely altered in neurons derived from alsin knockout (ALS2-/-) mice. We found that the Rab5-mediated endosome fusion was significantly increased in ALS2-/- neurons, resulting in an obvious decrease of endosome motility and increase of endosomes conversed to lysosomes in these neurons. Consequently, a significant increase of endosome/lysosome-dependent degradation of internalized cargo proteins such as glutamate receptors was observed in ALS2-/- neurons. Together, our data reveal a novel function of alsin in endosome trafficking, suggesting that the increased endosome degradation in ALS2-/- neurons underlies the pathogenic mechanism of ALS2 and related motor neuron diseases. This work is ready to submit to PNAS.